In recent years, techniques have been developed for analyzing single nucleotide polymorphisms (SNP) which have been shown to be associated with various diseases and drug side effects; in the development thereof, it is an important factor to accurately detect single nucleotide polymorphisms simply and in a short time.
An RFLP (Restriction Fragment Length Polymorphism) method is known as a method for analyzing single nucleotide polymorphisms. The RFLP method involves, when a restriction enzyme exists recognizing a gene mutation site in a PCR (Polymerase Chain Reaction) amplification product, preparing primers in common sequence sites, performing amplification by holding polymorphisms in the PCR amplification product, cleaving the resultant PCR product with the restriction enzyme, and determining the presence of polymorphisms based on the length of the fragments. However, the method has problems including that the use of restriction enzyme increases analysis cost and prolongs time of the whole analysis. It also has problems including that the detection of the chain length difference by electrophoresis complicates operation and prolongs time of the whole analysis.
In the fields of biochemistry, medicine, and the like, ion-exchange chromatography is used for the separation of biomacromolecules such as nucleic acids, proteins, and polysaccharides as a method capable of accurately detecting them simply and in a short time. The use of ion-exchange chromatography reduces complicated operation as required for measurement by electrophoresis. Non Patent Literature 1 discloses a method for separating nucleic acid-related compounds by high-performance liquid chromatography. However, even the method disclosed in Non Patent Literature 1 has a problem that it is difficult to sufficiently separate nucleic acids having chain lengths approaching to each other such as single nucleotide polymorphisms.